Vaccination with synthetic peptides corresponding to T-cell reactive fragments derived from tumor associated antigens (TAAs) can bolster anti-tumor immunity in patients expressing the same antigen. It has been increasingly acknowledged that TAA epitopes, in contrast to epitopes of viral origin, generally bind weakly to the MHC molecule and are therefore unable to elicit robust T cell response in vivo. Improving MHC binding as a strategy to upregulate antigen recognition has clear implications for immune intervention of cancer. Advances in the understanding of MHC/peptide interactions at the structural level over the past two decades have allowed the possibility of identifying novel binding motifs based on common conformational features. Analyses of the A2 molecule suggest a water-filled MHC/peptide binding interface. A direct implication of these studies is that hydrophilic amino acids in the central part of the bound peptide can indirectly interact with the MHC molecule and may modulate interactions with the T cell receptor. An Experimental Plan with two aims is proposed to: (i) study the pairwise interaction between two central residues in the MAGE-A4 epitope GVYDGREHTV on the diversity of TCR Vbeta CDR3 usage. (ii) derive and apply a common structural feature seen in eighteen x-ray structures to design of epitopes derived from Melan-A/MART-1, Her-2/neu, MUC1, CEA, MAGE-3, and AFP. As an outcome of the research proposed here, we expect to identify novel peptide sequences that are suitable as vaccine candidates in tumor immunotherapy. The research proposed in this application is significant, because upon successful completion, it will provide new criteria for converting low affinity TAA epitopes into high affinity variants that may recruit additional antigen-specific CTLs in vivo. By defining additional levels of sophistication of MHC/peptide/TCR interactions, more accurate predictions of immunogenic peptides will be possible. The research proposed will also benefit the development of peptide-based vaccines for viral and microbial infections.